Approximately one-third of veterans are current smokers, which is a significantly greater proportion than in the general population. Tobacco smoke exposure can promote airway hyperresponsiveness (AHR), a characteristic feature of asthma defined as airway constriction after exposure to a provocational agent. The prevalence of asthma is greater in veterans than in civilians and contributes significantly to healthcare utilization. Tobacco smoke is known to trigger asthma symptoms such as wheezing and cough; both are manifestations of airway hyperresponsiveness (AHR). Understanding the mechanisms behind AHR is important for patients that continue to be exposed to tobacco smoke (directly or secondhand), particularly asthmatics. Identifying a specific causative agent(s) for the adverse effects of smoking can be difficult given the complex nature of tobacco smoke. However, nicotine, a lipophilic chemical that is the major addictive component of tobacco smoke, may play an important role mediating the effects of tobacco smoke in the lung. Over the past several years, several studies have implicated nicotine in altered lung development and lung carcinogenesis. Nicotine is a ligand for endogenous nicotinic acetylcholine receptors (nAChRs). nAChRs are ligand gated channels comprised of five subunits that have neuronal and non-neuronal functions. The most common non neuronal receptor is the (17)5 nAChR. In this proposal, we hypothesize that chronic nicotine exposure promotes AHR by inducing nerve growth factor (NGF) secretion in lung fibroblasts through 17 nAChR dependent signaling. NGF belongs to the neurotrophin family, growth factors essential for development and survival of neurons. Overexpression of NGF has previously been associated with increased airway innervation and increased AHR after allergic stimulation. To address our hypothesis, we plan to utilize cell culture, organ explants, and a murine model of nicotine exposure. In Aim 1, using a cell culture model of primary lung fibroblasts and cultured lung explants from wild type and 17 nAChR deficient mice, we will demonstrate that nicotine acts on nAChRs to increase NGF secretion/production and increase neuronal growth. In Aim 2, we will demonstrate that airway smooth muscle contraction is increased and mediated by nerve growth factor by measuring bronchial ring contractility using isometric tension in an organ bath system. Bronchial rings will be harvested from unexposed mice and mice exposed to chronic nicotine. In Aim 3, we plan to use an in vivo mouse model of chronic nicotine exposure to show that NGF-related signaling is essential for nicotine induced increases in airway hyperresponsiveness by modifying the effects of nicotine with administration of NGF receptor agonists and antagonists. Airway hyperresponsiveness will be measured using a methacholine challenge in anesthetized, mechanically ventilated mice with the flexiVent. As part of this CDA-2 application, the applicant will engage in coursework geared toward improving grant/manuscript writing skills, and improving experimental design and analysis. She will also gain expertise in additional experimental techniques such as siRNA and isometric tension measurements. The applicant will also attend faculty development courses offered by her institution as well as national associations/professional societies that will address leadership, communication, and promotion. She will attend scientific meetings to present her research. These developmental activities will help the applicant achieve her immediate goals of improving her publication productivity and applying new research techniques, as well as her long term goal of becoming an independent investigator and mentor. With successful completion of the proposed project, we will better understand the mechanisms behind airway hyperresponsiveness related to nicotine exposure and help define alternative ways to treat veterans with asthma, particularly those who are exposed to tobacco smoke. PUBLIC HEALTH RELEVANCE: Cigarette smoking is more prevalent in veterans than the general population (35% vs. 21%). Exposure to tobacco smoke is also associated with airway hyperresponsiveness (AHR), a characteristic feature of asthma. Asthma is a common self reported diagnosis in veterans and contributes significantly to healthcare utilization. In asthmatics, cigarette smoke exposure is often a trigger of cough and wheezing, both manifestations of AHR. Given the increased prevalence of smoking and asthma in veterans, it is important to understand the mechanisms behind cigarette smoke associated AHR. Nicotine, the addictive component of cigarettes, may play an important role in AHR through neurogenic mechanisms. This project is relevant to the VA population because it will improve current understanding of the neurogenic pathways involved in nicotine exposure and may uncover alternative therapeutic interventions to improve disease management for asthmatic veterans, including those who continue to be exposed to cigarette smoke.